Protective Effect Of Hypoxic Preconditioning Research Articles

Hypoxic Preconditioning Averts Sporadic Alzheimer's Disease-Like Phenotype in Rats: A Focus on Mitochondria.

Aims: Brief episodes of sublethal hypoxia reprogram brain response to face possible subsequent lethal stimuli by triggering adaptive and prosurvival events-a phenomenon denominated hypoxic preconditioning (HP). To date, the potential therapeutic implications of HP to forestall sporadic Alzheimer's disease (sAD) pathology remain unexplored. Using a well-established protocol of HP and focusing on hippocampus as a first brain region affected in AD, this study was undertaken to investigate the potential protective effects of HP in a sAD rat model induced by the intracerebroventricular (icv) administration of streptozotocin (STZ) and to uncover the mitochondrial adaptations underlying this nonpharmacological strategy. Results: HP prevented the memory and learning deficits as well as tau pathology in the icvSTZ rat model. HP also attenuated icvSTZ-related reactive astrogliosis, as noted by increased glial fibrillary acidic protein immunoreactivity and myo-inositol levels. Notably, HP abrogated the icvSTZ-related impaired energy metabolism and oxidative damage. Particularly, HP averted increased lactate, glutamate, and succinate levels, and decreased mitochondrial respiratory chain function and mitochondrial DNA content. Concerning mitochondrial adaptations underlying HP-triggered tolerance to icvSTZ, preconditioned hippocampal mitochondria displayed an enhanced complex II-energized mitochondrial respiration, which resulted from a coordinated interaction between mitochondrial biogenesis and fusion-fission. Mitochondrial biogenesis was stimulated immediately after HP, whereas in a latter phase mitochondrial fusion-fission events are modulated favoring the generation of elongated mitochondria. Innovation and Conclusion: Overall, these results demonstrate for the first time that HP prevents the sAD-like phenotype, in part, by targeting mitochondria emerging as a preventive strategy in the context of AD. Antioxid. Redox Signal. 37, 739-757.

Attenuation of the cytoprotection induced by hypoxic preconditioning upon transfection with BNIP3-siRNA in human neuroblastoma SH-SY5Y cells

Purpose The aim of this study was to investigate the functional role of hypoxic preconditioning (HPC) in human neuroblastoma cells. Methods BNIP3 small-interfering RNA (BNIP3-siRNA) sequence was synthesized and used to transfect human neuroblastoma SH-SY5Y cell lines. Thereafter, BNIP3 expression at mRNA and protein levels and its effects on the cell proliferation were analyzed. The most effective pair of siRNA was selected to knockdown the expression level of BNIP3. Moreover, the effects of HPC on oxygen-glucose deprivation/reperfusion (OGD/R)-induced apoptosis and autophagy in SH-SY5Y cells were explored to further reveal the possible mechanisms underlying HPC. Results BNIP3-siRNA attenuated the protective effects of HPC by decreasing the cell viability, increasing the enzymatic activity of caspase-3 and 9, increasing the rate of apoptosis, and increasing the protein expression level of activated caspase-3. Additionally, BNIP3-siRNA had no significant influence on the expression level of HIF-1α induced by HPC, while it substantially inhibited HPC-induced BNIP3/Beclin1 and autophagy. Conclusions HPC promoted autophagy through regulating BNIP3 to reduce OGD/R.

SIRT1 mediates hypoxic preconditioning induced attenuation of neurovascular dysfunction following subarachnoid hemorrhage

Background and purposeVasospasm and delayed cerebral ischemia (DCI) contribute significantly to the morbidity/mortality associated with aneurysmal subarachnoid hemorrhage (SAH). While considerable research effort has focused on preventing or reversing vasospasm, SAH-induced brain injury occurs in response to a multitude of concomitantly acting pathophysiologic mechanisms. In this regard, the pleiotropic epigenetic responses to conditioning-based therapeutics may provide an ideal SAH therapeutic strategy. We previously documented the ability of hypoxic preconditioning (PC) to attenuate vasospasm and neurological deficits after SAH, in a manner that depends on the activity of endothelial nitric oxide synthase. The present study was undertaken to elucidate whether the NAD-dependent protein deacetylase sirtuin isoform SIRT1 is an upstream mediator of hypoxic PC-induced protection, and to assess the efficacy of the SIRT1-activating polyphenol Resveratrol as a pharmacologic preconditioning therapy. MethodsWild-type C57BL/6J mice were utilized in the study and subjected to normoxia or hypoxic PC. Surgical procedures included induction of SAH via endovascular perforation or sham surgery. Multiple endpoints were assessed including cerebral vasospasm, neurobehavioral deficits, SIRT1 expression via quantitative real-time PCR for mRNA, and western blot for protein quantification. Pharmacological agents utilized in the study include EX-527 (SIRT1 inhibitor), and Resveratrol (SIRT1 activator). ResultsHypoxic PC leads to rapid and sustained increase in cerebral SIRT1 mRNA and protein expression. SIRT1 inhibition blocks the protective effects of hypoxic PC on vasospasm and neurological deficits. Resveratrol pretreatment dose-dependently abrogates vasospasm and attenuates neurological deficits following SAH – beneficial effects that were similarly blocked by pharmacologic inhibition of SIRT1. ConclusionSIRT1 mediates hypoxic preconditioning-induced protection against neurovascular dysfunction after SAH. Resveratrol mimics this neurovascular protection, at least in part, via SIRT1. Activation of SIRT1 is a promising, novel, pleiotropic therapeutic strategy to combat DCI after SAH.

BDNF Overexpression Enhances the Preconditioning Effect of Brief Episodes of Hypoxia, Promoting Survival of GABAergic Neurons

Hypoxia causes depression of synaptic plasticity, hyperexcitation of neuronal networks, and the death of specific populations of neurons. However, brief episodes of hypoxia can promote the adaptation of cells. Hypoxic preconditioning is well manifested in glutamatergic neurons, while this adaptive mechanism is virtually suppressed in GABAergic neurons. Here, we show that brain-derived neurotrophic factor (BDNF) overexpression in neurons enhances the preconditioning effect of brief episodes of hypoxia. The amplitudes of the NMDAR- and AMPAR-mediated Ca2+ responses of glutamatergic and GABAergic neurons gradually decreased after repetitive brief hypoxia/reoxygenation cycles in cell cultures transduced with the (AAV)-Syn-BDNF-EGFP virus construct. In contrast, the amplitudes of the responses of GABAergic neurons increased in non-transduced cultures after preconditioning. The decrease of the amplitudes in GABAergic neurons indicated the activation of mechanisms of hypoxic preconditioning. Preconditioning suppressed apoptotic or necrotic cell death. This effect was most pronounced in cultures with BDNF overexpression. Knockdown of BDNF abolished the effect of preconditioning and promoted the death of GABAergic neurons. Moreover, the expression of the anti-apoptotic genes Stat3, Socs3, and Bcl-xl substantially increased 24 h after hypoxic episodes in the transduced cultures compared to controls. The expression of genes encoding the pro-inflammatory cytokines IL-10 and IL-6 also increased. In turn, the expression of pro-apoptotic (Bax, Casp-3, and Fas) and pro-inflammatory (IL-1β and TNFα) genes decreased after hypoxic episodes in cultures with BDNF overexpression. Inhibition of vesicular BDNF release abolished its protective action targeting inhibition of the oxygen-glucose deprivation (OGD)-induced [Ca2+]i increase in GABAergic and glutamatergic neurons, thus promoting their death. Bafilomycin A1, Brefeldin A, and tetanus toxin suppressed vesicular release (including BDNF) and shifted the gene expression profile towards excitotoxicity, inflammation, and apoptosis. These inhibitors of vesicular release abolished the protective effects of hypoxic preconditioning in glutamatergic neurons 24 h after hypoxia/reoxygenation cycles. This finding indicates a significant contribution of vesicular BDNF release to the development of the mechanisms of hypoxic preconditioning. Thus, our results demonstrate that BDNF plays a pivotal role in the activation and enhancement of the preconditioning effect of brief episodes of hypoxia and promotes tolerance of the most vulnerable populations of GABAergic neurons to hypoxia/ischemia.

Priming of the Cells: Hypoxic Preconditioning for Stem Cell Therapy.

Objective:Stem cell-based therapies are promising in regenerative medicine for protecting and repairing damaged brain tissues after injury or in the context of chronic diseases. Hypoxia can induce physiological and pathological responses. A hypoxic insult might act as a double-edged sword, it induces cell death and brain damage, but on the other hand, sublethal hypoxia can trigger an adaptation response called hypoxic preconditioning or hypoxic tolerance that is of immense importance for the survival of cells and tissues.Data Sources:This review was based on articles published in PubMed databases up to August 16, 2017, with the following keywords: “stem cells,” “hypoxic preconditioning,” “ischemic preconditioning,” and “cell transplantation.”Study Selection:Original articles and critical reviews on the topics were selected.Results:Hypoxic preconditioning has been investigated as a primary endogenous protective mechanism and possible treatment against ischemic injuries. Many cellular and molecular mechanisms underlying the protective effects of hypoxic preconditioning have been identified.Conclusions:In cell transplantation therapy, hypoxic pretreatment of stem cells and neural progenitors markedly increases the survival and regenerative capabilities of these cells in the host environment, leading to enhanced therapeutic effects in various disease models. Regenerative treatments can mobilize endogenous stem cells for neurogenesis and angiogenesis in the adult brain. Furthermore, transplantation of stem cells/neural progenitors achieves therapeutic benefits via cell replacement and/or increased trophic support. Combinatorial approaches of cell-based therapy with additional strategies such as neuroprotective protocols, anti-inflammatory treatment, and rehabilitation therapy can significantly improve therapeutic benefits. In this review, we will discuss the recent progress regarding cell types and applications in regenerative medicine as well as future applications.

The key role of microtubules in hypoxia preconditioning-induced nuclear translocation of HIF-1α in rat cardiomyocytes.

BackgroundHypoxia-inducible factor (HIF)-1 is involved in the regulation of hypoxic preconditioning in cardiomyocytes. Under hypoxic conditions, HIF-1α accumulates and is translocated to the nucleus, where it forms an active complex with HIF-1β and activates transcription of approximately 60 kinds of hypoxia-adaptive genes. Microtubules are hollow tubular structures in the cell that maintain cellular morphology and that transport substances. This study attempted to clarify the role of microtubule structure in the endonuclear aggregation of HIF-1α following hypoxic preconditioning of cardiomyocytes.MethodsPrimary rat cardiomyocytes were isolated and cultured. The cardiomyocyte culture system was used to establish a hypoxia model and a hypoxic preconditioning model. Interventions were performed on primary cardiomyocytes using a microtubule-depolymerizing agent and different concentrations of a microtubule stabilizer. The microtubule structure and the degree of HIF-1α nuclear aggregation were observed by confocal laser scanning microscopy. The expression of HIF-1α in the cytoplasm and nucleus was detected using Western blotting. Cardiomyocyte energy content, reflected by adenosine triphosphate/adenosine diphosphate (ATP/ADP), and key glycolytic enzymes were monitored by colorimetry and high-performance liquid chromatography (HPLC). Reactive oxygen species (ROS) production was also used to comprehensively assess whether microtubule stabilization can enhance the myocardial protective effect of hypoxic preconditioning.ResultsDuring prolonged hypoxia, it was found that the destruction of the microtubule network structure of cardiomyocytes was gradually aggravated. After this preconditioning, an abundance of HIF-1α was clustered in the nucleus. When the microtubules were depolymerized and hypoxia pretreatment was performed, HIF-1α clustering occurred around the nucleus, and HIF-1α nuclear expression was low. The levels of key glycolytic enzymes were significantly higher in the microtubule stabilizer group than in the hypoxia group. Additionally, the levels of lactate dehydrogenase and ROS were significantly lower in the microtubule stabilizer group than in the hypoxia group.ConclusionThe microtubules of cardiomyocytes may be involved in the process of HIF-1α endonuclear aggregation, helping to enhance the anti-hypoxic ability of cardiomyocytes.

Autophagy mediates the beneficial effect of hypoxic preconditioning on bone marrow mesenchymal stem cells for the therapy of myocardial infarction

BackgroundStem cell therapy has emerged as a promising therapeutic strategy for myocardial infarction (MI). However, the poor viability of transplanted stem cells hampers their therapeutic efficacy. Hypoxic preconditioning (HPC) can effectively promote the survival of stem cells. The aim of this study was to investigate whether HPC improved the functional survival of bone marrow mesenchymal stem cells (BM-MSCs) and increased their cardiac protective effect.MethodsBM-MSCs, isolated from Tg(Fluc-egfp) mice which constitutively express both firefly luciferase (Fluc) and enhanced green fluorescent protein (eGFP), were preconditioned with HPC (1% O2) for 12 h, 24 h, 36 h, and 48 h, respectively, followed by 24 h of hypoxia and serum deprivation (H/SD) injury.ResultsHPC dose-dependently increased the autophagy in BM-MSCs. However, the protective effects of HPC for 24 h are most pronounced. Moreover, hypoxic preconditioned BM-MSCs (HPCMSCs) and nonhypoxic preconditioned BM-MSCs (NPCMSCs) were transplanted into infarcted hearts. Longitudinal in vivo bioluminescence imaging (BLI) and immunofluorescent staining revealed that HPC enhanced the survival of engrafted BM-MSCs. Furthermore, HPCMSCs significantly reduced fibrosis, decreased apoptotic cardiomyocytes, and preserved heart function. However, the beneficial effect of HPC was abolished by autophagy inhibition with 3-methyladenine (3-MA) and Atg7siRNA.ConclusionThis study demonstrates that HPC may improve the functional survival and the therapeutic efficiencies of engrafted BM-MSCs, at least in part through autophagy regulation. Hypoxic preconditioning may serve as a promising strategy for optimizing cell-based cardiac regenerative therapy.

Proteomic analysis of the mouse brain after repetitive exposure to hypoxia

Hypoxic preconditioning (HPC) is known to have a protective effect against hypoxic damage; however, the precise mechanisms involved remain unknown. In this study, an acute and repetitive hypoxia mouse model, two-dimensional fluorescence difference gel electrophoresis (2D-DIGE) coupled with matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF/TOF-MS), and Western blot experiments were used to identify the differential expression of key proteins in the mouse brain during HPC. Approximately 2100 2D-DIGE spots were observed following gel imaging and spot detection. Significant differences (p<0.05) in the expression of 66 proteins were observed between the 3× HPC treatment group and the control group, 45 proteins were observed between the 6× HPC treatment group and the control group, and 70 proteins were observed between the 3× HPC treatment group and the 6× HPC group. Consistent results among Western blot, 2D-DIGE and MS methods were observed for the proteins, ATP synthase subunit alpha, malate dehydrogenase, guanine nucleotide-binding protein subunit beta-1 and proteasome subunit alpha type-2. The proteins associated with ATP synthesis and the citric acid cycle were down-regulated, while those linked to glycolysis and oxygen-binding were up-regulated. This proteomic analysis of the mouse brain after HPC furthers understanding of the molecular pathways involved in the protective effect of HPC and these findings provide new insight into the mechanisms of hypoxia and HPC.

Myocardial protective effect of hypoxic preconditioning with double cardiopulmonary bypass in canine hearts

Objective To investigate the myocardial protective effect of hypoxic preconditioning in thein vivo canine hearts.Methods Healthy beagle dogs were randomly divided into two groups:the hypoxic preconditioning group (HPC group,n =8) and the control group (C group,n =8).The dog model of the double cardiopulmonary bypass (systemic and coronary circulation) was established after chest opening under general anesthesia.In HPC group,dog hearts were perfused by the coronary circulation and subjected to hypoxic preconditioning (3 min of hypoxia followed by 5 min of reoxygenation,repeated 3 times).The animals in C group were not subjected to hypoxic preconditioning.Both groups underwent cold cardioplegia perfusion through coronary circulation to induce cardiac arrest.At 60 min after cardiopulmonary bypass,we observed one-time rebeat rate,the incidence of arrhythmia and electric defibrillation times.At 30 min after rebeating,the left anterior descending coronary arteries were ligated with suture at the lower 1/3.The myocardial infarction area size was observed 60 min later,and samples were taken for examination by electron microscopy.At T1 (before operation),T2 (before cardiopulmonary bypass),T3 (immediately after the rebeat),and T4 (30 min after the ligation of left anterior descending coronary),the troponin and myocardial enzymes spectrum were measured.Cardiac function and myocardial structure were also observed in both groups.Results Myocardial enzymes at T3 were higher in HPC group than in C group (P ＜ 0.05),and myocardial enzymes and troponin at T4 were significantly lower in HPC group (P ＜ 0.05).As compared with C group,one-time rebeat rate and defibrillation times were significantly reduced in HPC group (7/8 vs.4/8 and 1/8 vs.5/8,P＜0.05).The infarct area size was significantly smaller in HPC group than in C group [(15.37 ± 1.64) ％ vs.(29.77 ± 3.38) ％,P ＜ 0.05],and electron microscopy revealed that the damage of mitochondrial structure and edema of cardiomyocytes were significantly reduced in HPC group as compared with C group.Conclusion Hypoxia preconditioning of the whole heart in vivo had protective effect on myocardia.It could improve the tolerance of myocardia to hypoxic injury. Key words: Hypoxic preconditioning; Cardiopulmonary bypass; Myocardial protection

HYPOXIC PRECONDITIONING OF STEM CELLS AS A NEW APPROACH TO INCREASE THE EFFICACY OF CELL THERAPY FOR MYOCARDIAL INFARCTION

During the last decade, stem cell research has developed at an accelerated pace. Various types of stem cells have been tested for myocardial infarction therapy. Despite the preclinical benefits of cell therapy success in clinical trials remains modest. The main obstacles to regeneration of the infarcted heart using stem cells are: 1) not every stem cell type can differentiate into cardiomyocytes; and 2) low survival rates of transplanted cells, due to the harsh environment of the infarcted myocardium. Hypoxic preconditioning (HP) has been shown to improve transplantation efficacy of mesenchymal stem cells and cardiac progenitor cells in animal models of myocardial infarction. It has also been shown that transplantation of preconditioned cells decreases infarct size, prevents postinfarction remodeling of the heart, and positively modulates development of ischemic cardiomyopathy. Hypoxic preconditioning also prevents extensive death of transplanted cells due to necrosis and apoptosis during long-term hypoxia or oxidative stress. The protective effect of HP is based on three main processes: (1) modification of cell phenotypes to help survival during hypoxia (enhancement of HIF-1alpha expression, ERK1/2 and Akt activation, enhancement of erythropoietin receptor expression and erythropoietin production, and an elevation in levels of antiapoptotic proteins Bcl-2 and Bcl-xL); (2) upregulation of various secretable factors including the vascular endothelial growth factor (VEGF) and hepatocyte growth factor (HGF), and expression of VEGF-2 and HGF-receptors; (3) enhancement in the formation of CXCR4 and CXCR7 receptors, which play an important role in mobilization and homing of stem cells in the ischemic region.

Combination of Hypoxic Preconditioning and Postconditioning Does Not Induce Additive Protection of Ex Vivo Human Skeletal Muscle From Hypoxia/Reoxygenation Injury

We previously demonstrated that hypoxic preconditioning (HPreC) or postconditioning (HPostC) protected ex vivo human skeletal muscle from hypoxia/reoxygenation injury. Here, we investigated if combined HPreC and HPostC could convey additive protection. Human rectus abdominis muscle strips were cultured in normoxic Krebs buffer for 5 hours (control) or in 3 hours hypoxic/2 hours normoxic buffer (treatment groups). HPreC and HPostC were induced by 1 cycle of 5 minutes hypoxia/5 minutes reoxygenation immediately before or after 3 hours hypoxia, respectively. Muscle injury, viability, and adenosine triphosphate (ATP) synthesis were assessed by measuring lactate dehydrogenase release, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide reduction, and ATP content, respectively. Hypoxia/reoxygenation caused lactate dehydrogenase to increase and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide reduction and ATP content to decrease (P < 0.05; n = 7). HPreC, HPostC, and combination of both were equally effective in protection of muscle from hypoxia/reoxygenation injury. Atractyloside (5 × 10 M), a mitochondrial permeability transition pore opener, abolished the protective effect of HPreC or HPostC. We conclude that HPreC and HPostC protect ex vivo human skeletal muscle against hypoxia/reoxygenation injury by closing the mitochondrial permeability transition pore. For that reason, they are equally effective and do not demonstrate an additive effect. Moreover, the potent effect of HPostC indicates ischemic postconditioning as an effective clinical intervention against reperfusion injury in autogenous skeletal muscle transplantation and replantation surgery.

Peroxisome proliferator activated receptor (PPAR)-γ co-activator 1-α and hypoxia induced factor-1α mediate neuro- and vascular protection by hypoxic preconditioning in vitro

Preconditioning-induced cellular adaptation is a new therapeutic strategy for ischemic stroke. This research aims to examine the role of peroxisome proliferator activated receptor (PPAR)-γ co-activator 1-α (PGC-1α) and hypoxia induced factor-1α (HIF-1α) in hypoxic preconditioning-induced protection. In this study, rat artery endothelial cells and neuronal PC12 cells were preconditioned with hypoxia before oxygen–glucose deprivation (OGD) insult. Cell viability, protein expression and oxidative stress were then evaluated. PGC-1α and HIF-1α were knocked down by RNA interference. We found that hypoxic preconditioning significantly reduced cell damage, enhanced the expression of PGC-1α, HIF-1α and VEGF and attenuated oxidative stress in endothelial and PC12 cells in OGD model. The protective effects of hypoxic preconditioning were hardly detected in HIF-1α or PGC-1α deficit cells. The loss of protection was accompanied with a significant loss of VEGF expression in HIF-1α or PGC-1α deficit PC12 cells and PGC-1α deficit endothelial cells as well as a considerable decrease of anti-oxidative effects in PGC-1α knocked-down endothelial cells. The present study demonstrated that both PGC-1α and HIF-1α played crucial roles in hypoxic preconditioning in endothelial and neuronal cells.

Hypoxic Preconditioning Enhances Survival of Human Adipose-Derived Stem Cells and Conditions Endothelial Cells In Vitro

To grow more robust cardiac tissue for implantation in vivo, strategies to improve survival of implanted stem cells are required. Here we report the protective effects of hypoxic preconditioning (HPC) and identify mechanisms for improving survival of adipose-derived stem cells (ASC) in vitro. Human ASC were preconditioned for 24 h with hypoxia and then exposed to simulated ischemia for a further 24 h. HPC significantly increased ASC viability, and reduced cell injury and apoptosis compared with non-preconditioned cells under ischemic conditions, as shown by 3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT), lactate dehydrogenase-release, and caspase activity assays. Preconditioned ASC increased levels of hypoxia-inducible factor-1 alpha and secreted significantly more of the downstream target vascular endothelial growth factor (VEGF-A; 13-fold) compared with control during the 24 h. Exogenous VEGF (50 ng/mL) increased phosphorylation of Akt without affecting ERK1/2, JNK, or p38 MAPK protein levels. Phospho-Akt was also increased in preconditioned ASC compared with non-preconditioned ASC, an effect that may be mediated via VEGF-A. Importantly, the protective effects of HPC were abolished by a neutralizing antibody against VEGF-A and the phosphoinositol 3-kinase inhibitor LY294002, demonstrating the importance of VEGF-A and Akt in hypoxia-induced ASC survival. Importantly, we showed that media derived from hypoxic preconditioned ASC support endothelial cell survival and endothelial tube formation in vitro. Our in vitro findings indicate that HPC may be a promising strategy to improve survival of ASC and promote angiogenesis in ischemic environments.

Involvement of the transcription factor c-Fos in the protective effect of hypoxic preconditioning in a model of post-traumatic stress disorder in rats

A quantitative immunocytochemical method was used to study the dynamics of the expression of the transcription factor c-Fos in the hypothalamus, hippocampus, and neocortex during the development of anxiety in the post-traumatic stress disorder (PTSD) model in control rats and rats preconditioned by hypoxia, which is known to have an anxiolytic effect. It was found that the induction of experimental PTSD was associated with a significant and stable increase in the level of c-Fos in all investigated brain areas. The maximum increase according to amplitude (up to 30 times) and duration (about 10 days) was observed in the hypothalamus. This overexpression may be associated with corticotropin-releasing hormone hyperproduction, which is typical of PTSD. In animals that were subjected to triple hypoxic preconditioning with mild repetitive hypobaric hypoxia (360 mm Hg for 2 hours daily, for 3 days), which prevented the traumatic stress-induced pathology, c-Fos overexpression was completely or partially blocked. These data indicate that stable overexpression of the c-Fos transcription factor in the neocortex, hippocampus, and hypothalamus apparently participates in the mechanisms of PTSD development, whereas its blockade plays an important role in increasing the brain’s tolerance to stresses and in protection against stress-induced pathologies.

Hypoxic preconditioning induces neuroprotection against transient global ischemia in adult rats via preserving the activity of Na +/K +-ATPase

We demonstrated previously that 30 min of hypoxic preconditioning (HPC) applied 1 day before 10 min of transient global cerebral ischemia (tGCI) reduced neuronal loss in the hippocampal CA1 subregion in adult rats. The aim of the present study was to investigate the role of Na +/K +-ATPase and protein kinase Mζ (PKMζ) in the protective effect of HPC against tGCI in adult rats. We found that the activity of Na +/K +-ATPase decreased in the hippocampal CA1 subregion after 10 min of tGCI. This effect was not seen after 30 min of HPC in adult rats. Corresponding to the changes in Na +/K +-ATPase activity, the surface expression of Na +/K +-ATPase α1 subunit increased after HPC. Furthermore, HPC dramatically reduced the number of terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL)-positive cells in the hippocampal CA1 subregion after tGCI. However, neither PKMζ nor phosphorylation of PKMζ was changed after tGCI or HPC. The results of the present study are consistent with the hypothesis that both enhanced recovery of Na +/K +-ATPase activity due to preserved the protein levels of Na +/K +-ATPase α1 subunit and reduced DNA fragmentation after tGCI contribute to the protection afforded by HPC. However, PKMζ activation does not appear to play a role in this neuroprotection.

Protective Effect of Hypoxic Preconditioning on Hypoxic-Ischemic Injured Newborn Rats

Brief episodes of cerebral hypoxia-ischemia cause transient ischemic tolerance to subsequent ischemic events that are otherwise lethal. This study was conducted to evaluate the protective effect of hypoxic preconditioning on hypoxic-ischemic injury in the neonatal rat and the persistence of a protective window after hypoxic preconditioning. The rats were preconditioned with hypoxia (8% oxygen, 92% nitrogen) for three hours, subjected to ischemia using ligation of the right common carotid artery, and then exposed to another three hours of hypoxia. Using proton magnetic resonance spectroscopy, terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end-labeling (TUNEL) staining, and morphologic scores, this study shows that hypoxic preconditioning 6-hr to 1-day before hypoxic-ischemic injury increases survival rates and has neuroprotective effects against subsequent hypoxic-ischemic injury. The mechanism of the protective effects of hypoxic preconditioning in the newborn rat brain may involve downregulation of apoptotic cell death.

Activation of the Central Histaminergic System is Involved in Hypoxia-Induced Stroke Tolerance in Adult Mice

We hypothesized that activation of the central histaminergic system is required for neuroprotection induced by hypoxic preconditioning. Wild-type (WT) and histidine decarboxylase knockout (HDC-KO) mice were preconditioned by 3 hours of hypoxia (8% O(2)) and, 48 hours later, subjected to 30 minutes of middle cerebral artery (MCA) occlusion, followed by 24 hours of reperfusion. Hypoxic preconditioning improved neurologic function and decreased infarct volume in WT or HDC-KO mice treated with histamine, but not in HDC-KO or WT mice treated with α-fluoromethylhistidine (α-FMH, an inhibitor of HDC). Laser-Doppler flowmetry analysis showed that hypoxic preconditioning ameliorated cerebral blood flow (CBF) in the periphery of the MCA territory during ischemia in WT mice but not in HDC-KO mice. Histamine decreased in the cortex of WT mice after 2, 3, and 4 hours of hypoxia, and HDC activity increased after 3 hours of hypoxia. Vascular endothelial growth factor (VEGF) mRNA and protein expressions showed a greater increase after hypoxia than those in HDC-KO or α-FMH-treated WT mice. In addition, the VEGF receptor-2 antagonist SU1498 prevented the protective effect of hypoxic preconditioning in infarct volume and reversed increased peripheral CBF in WT mice. Therefore, endogenous histamine is an essential mediator of hypoxic preconditioning. It may function by enhancing hypoxia-induced VEGF expression.

Influence of hypoxic-preconditioning on autonomic regulation following global ischemic brain injury in rats

Experimental and clinical studies have shown that autonomic imbalance is associated with morbidity and mortality due to global ischemic brain injury following cardiac arrest (CA). Although hypoxic-preconditioning (HP) has shown promising neuro-protection in the subsequent ischemic brain injury, the underlying mechanisms and its influence on autonomic regulation have not yet well-understood. In this study, we utilized baroreflex sensitivity (BRS) to investigate the protective effect of HP on autonomic regulation. We investigated changes in heart rate, arterial blood pressure (BP), and BRS within 4 h after CA in rats. The relationship between BRS and neurodeficit score (NDS) was analyzed. Although no significant differences were found in heart rate and BP before and after CA between the control and the preconditioned groups, both BRS and NDS of preconditioned rats were clearly higher than that of the control rats during recovery after CA. Furthermore, BRS in the first 4 h after CA highly correlated with NDS 24 h after CA. These results imply that treatment with HP improves autonomic regulation and protects the brain from ischemic injury. The correlation between BRS and NDS also suggests that BRS can be a prognostic criterion for the level of brain injury after CA.

Hypoxia interferes with connective tissue growth factor (CTGF) gene expression in human proximal tubular cell lines

Hypoxia plays an important role in kidney injury. By the stabilization of the transcription factor HIF-1, hypoxia affects gene expression also in tubular epithelial cells. Increased expression of connective tissue growth factor (CTGF) is observed in different kidney diseases and is associated with deteriorating renal function. Therefore, we hypothesized that the expression of CTGF might be modulated under hypoxic conditions. The human proximal tubular epithelial cell lines HK-2 and HKC-8 were treated with reduced oxygen tension (1% O(2)) or the hypoxia mimetic dimethyloxalyl glycine (DMOG). CTGF was analysed by Western blotting, real-time RT-PCR and luciferase gene expression assays. Exposure of HK-2 or HKC-8 cells to hypoxia or treatment with DMOG for up to 24 h reduced cellular as well as secreted CTGF protein synthesis. Downregulation was also detectable at the mRNA level and was confirmed by reporter gene assays. Hypoxic repression of CTGF synthesis was dependent on HIF-1, as shown by HIF-1alpha knockdown by siRNA. Furthermore, exposure to hypoxia reduced CTGF synthesis in response to TGF-beta. A negative correlation between HIF-1alpha accumulation and CTGF synthesis was also observed in renal cell carcinoma cells (RCC4 and RCC10). Reexpression of von Hippel-Lindau protein reduced HIF-1alpha and increased CTGF synthesis. We provide evidence that hypoxia inhibits CTGF synthesis in human proximal tubular epithelial cells, involving HIF-1alpha. Under hypoxic conditions, induction of CTGF by TGF-beta was repressed. The reduced synthesis of the profibrotic factor CTGF may contribute to a potential protective effect of hypoxic preconditioning in acute renal injury.

Hypoxia-inducible factor-1α has a key role in hypoxic preconditioning

Sublethal hypoxia induces tolerance to subsequent hypoxic insults in a process known as hypoxic preconditioning (HP). Hypoxia-inducible factor-1α (HIF-1α) is a key transcription protein involved in the mechanism of HP. In this study, we investigated the effects of HP on tissue oxygenation and expression of HIF-1α gene targets in the brain using neural cell-specific HIF-1α-deficient mice. The animals were exposed to 8% oxygen for 3 hours. Twenty-four hours later, the oxygen partial pressure (pO 2) of brain tissue and gene expression were measured during hypoxia. HP improved the pO 2 of brain tissue during subsequent hypoxia with upregulated inducible nitric oxide synthase in wild-type mice, whereas HP had no detectable effect in the mutant mice. Our results indicate that the protective effects of HP may be partially mediated by improving tissue oxygenation via HIF-1α and inducible nitric oxide synthase.